The broad, long-term objective of this project is to ascertain whether alterations in 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] synthesis or in the molecular action of this steroid hormone provide the basis of vitamin D- dependent forms of absorptive hypercalciuria (AH). Specifically, we will focus on the biochemistry and molecular biology of the vitamin D receptor (VDR) to search for the cause of increased intestinal calcium absorption invariably found in AH. This objective will be approached by 6 specific aims. Aim 1 will explore the biochemistry of the VDR in skin fibroblasts (number, affinity,) and function (24-OHase activity) from patients with AH. Aim 2 will also utilize studies in skin fibroblasts but will use molecular biological techniques to quantitate VDR mRNA, measure the sensitivity of the 1,25(OH)2D3-induced upregulation of VDR mRNA and assess VDR mRNA synthesis and stability. The hypothesis to be tested in these two aims is that VDR mRNA stability would be increased in primary receptor upregulation (due to abnormality in the VDR mRNA non-coding region) or in secondary receptor upregulation due to facilitated 1-OHase. Aim 3 will continue to explore the nature of the VDR allelic variation observed in' the 3'-noncoding region by ascertaining whether it is a marker for the disease and, if so, further examine its location and nature of the alteration. Additional studies will also assess whether similar polymorphisms may be found in the 5'-noncoding region. Aim 4 will begin to explore the activity of the 25-OHD3-1alpha-OHase using appropriate probes if and when they become available. Aim 5 will examine the effect of phosphate therapy on the biochemical and molecular biological parameters outlined above. Aim 6 will explore the use of other classic vitamin D target cells for many of the above mentioned studies while serving to verify the observations made in fibroblasts. Overall, these specific aims should delineate the contribution of abnormalities in the VDR gene, or in the over-expression of this gene, to the development of AH.